Irrigation solution and methods for use

ABSTRACT

Methods and solutions are provided for removal of the smear layer on prepared tooth and bone surfaces, especially in endodontic environments.

This application is a Continuation in Part of U.S. application Ser. No.10/055,075, filed Jan. 23, 2002 now abandoned, which is hereinincorporated by reference in its entirety.

The present invention relates to methods and solutions for removingundesirable substances from tooth surfaces during dental procedures. Theinvention removes buildup of debris and bacteria formed duringpreparation of tooth surfaces during procedures such as root canaltreatment, restoration, dental reconstruction, periodontal procedures,and the like, and is also suitable for preparation of bone forreconstruction or restoration.

As a consequence of pathological changes in the dental pulp, the rootcanal system acquires the capacity to harbor several species ofbacteria, their toxins and their by-products. The microorganisms presentin infected root canals are predominantly gram-negative anaerobes thatare seeded into the root canals from direct pulp exposures (caries ortraumatic injuries) or coronal microleakage. The morphology of rootcanals is very complex and mechanically prepared root canals containareas that cannot be reached by endodontic instruments. Themicroorganisms present in the root canal not only invade the anatomicirregularities of the root canal system, but also invade the dentinaltubules.

In the root, dentinal tubules extend from the intermediate dentin justinside the cementum-dentin junction to the pulp-predentin junction.Tubules are approximately 1 μm in diameter near the cementum-dentinjunction and approximately 2.5 μm near the pulp-predentin junction. Thenumber of dentinal tubules per square millimeter varies from 8,000 to57,000. At the periphery of the root at the cemento-enamel junction, thenumber has been estimated to be approximately 15,000 per squaremillimeter.

Many studies have shown that currently used methods of cleaning andshaping produce a smear layer that covers root canal walls. The smearlayer is produced as a result of instrumentation and its content isforced into the dentinal tubules to varying distances. Moodnik, R. M.,Dorn, S. O., Feldman, M. J., Levey, M., and Borden, B. G., J. Endodon.,1976, 2, 261-266; Cengiz, T., Aktener, B. O., and Piskin, B., Int'l.Endodon. J., 1990, 23, 163-171. Cengiz, et al. suggested that thepenetration of smear material into the dentinal tubules is probablycaused by capillary action generated between the dentinal tubules andthe smear material.

In 1975, McComb and Smith described the smear layer in endodontics.McComb, D., and Smith, D. C., J. Endodon., 1975, 1, 238-242. It waslater characterized as consisting of a superficial layer on the surfaceof the canal wall that averages between 1-2 μm in thickness, and adeeper layer packed into the dentinal tubules to a depth of up to 40 μm.Cameron, J. A., J. Endodon., 1983, 9, 289-292; Mader, C. L.,Baumgartner, J. C., and Peters, D. D., J. Endodon., 1984, 10, 477-483.The smear layer consists of organic and inorganic substances thatinclude fragments of odontoblastic processes, microorganisms andnecrotic materials. A number of studies have shown that presence ofsmear layer can prevent penetration of root canal medications andsealers into the dental tubules. In addition, they have shown thatremoval of the smear layer results in better adaptation between rootcanal filling materials and the dentinal walls.

Bacteria present in the infected root canals usually invade the dentinaltubules and can re-infect the root canals if they remain viable afterroot canal therapy. Viable bacteria has been reported in dentinaltubules of infected teeth at approximately half the distance between theroot canal walls and the cemento-dentinal junction. Endotoxins have beenfound within the dentinal walls of infected root canals as well. Concernhas been evidenced regarding the fate of these bacteria, especiallywhether they may find nutrients for growth and reproduction.

Complete eradication of bacteria present in the canals and dentinaltubules, sealing root canals in three dimensions and prevention ofrecontamination of sealed root canals are the ideal goals for endodontictherapy. Because of the complexity of root canal systems, and theinability of instruments to contact all surfaces of the root canals, itis impossible to create a sterile space in all teeth with infected rootcanals. Bystrom, A., and Sundqvist, G., Scand, J. Dent. Res., 1981, 89,321-328; Bystrom, A., Claesson, R., and Sundqvist, G., Endod. Dent.Traumatol., 1985, 1, 170-175. In fact, residual bacteria in aninstrumented and unfilled canal can multiply to their original numberswithin 2-4 days. To prevent repopulation of the root canals withresidual bacteria, the use of intracanal medications and completion oftreatment of infected root canals in more than one visit has beenrecommended. Bystrom, A., Claesson, R., and Sundqvist, G., Endod. Dent.Traumatol., 1985, 1, 170-175; Chong, B. S., and Pitt Ford, T. R., Int'l.Endodon. J., 1992, 25, 97-106.

Intracanal medications have traditionally been considered important tosuccess of root canal therapy. In fact, it has been a common assumptionthat success, both short- and long-term, depends on the chemicals placedin the canal between appointments. However, there is no firm scientificevidence for usefulness of medications such as camphoratedmonochlorophenol (CMCP), formocresol, cresatin, or calcium hydroxide(Ca(OH)₂), which have been used as intracanal medications. Theintracanal medicaments have been proposed for, inter alia, antimicrobialuse in the pulp and periapex, neutralization of canal remnants to renderthem inert, and control or prevention of post-treatment pain.

A study of the presence and influence of bacteria on the long-termsuccess of root canal therapy showed that about 40% of root canals areinfected after instrumentation. Sjogren, U., Figdor, D., Persson, S.,and Sundqvist, G., Int'l; Endodon. J., 1997, 30, 297-306. In addition,instrumented canals without application of an intracanal medication withCa(OH)₂ failed significantly more frequently than those which weremedicated for one week with Ca(OH)₂ (68% vs. 94%). The results of thisstudy corroborate a 1987 study showing improved clinical success ratesfollowing effective disinfection of root canals. Bystrom, A., Happonen,R., Sjogren, U., and Sundqvist, G., Endod. Dent. Traumatol., 1987, 3,58-63.

According to a number of authorities, presence of smear layer caninhibit penetration of anti-microbial agents such as intra-canalirrigants and medicaments into the dentinal tubules. Haapasalo, M., andOrstavik, D., J. Dent. Res., 1987, 66, 1375-1379; Czonstkowsky, M.,Wilson, E., and Holstein, F., Dental Clinics of N. Am., 1990, 34, 13-24.Several investigators have reported better adhesion of obturationmaterials to the canal walls after removal of the smear layer. Goldberg,F., and Abramovich, A., J. Endodon., 1977, 3, 101-105; White, R. R.,Goldman, M., and Lin, P. S., J. Endodon., 1984, 10, 558-562. Severalstudies have also reported poor or no penetration of sealer in tubuleswith an intact smear layer. These studies have shown improvedpenetration following removal of the smear layer with sealers such asTubliseal (penetration to 15 μm); AH26 (penetration from 10-60 μm); andSealpex, Roth's 811, and CRCS (all with penetration from 35-80 μm).Gutierrez, J. H., Herrera, V. R., Berg, E. H., Villena, F., and Jofre,A., Oral Surg. Oral Med. Oral Path., 1990. 70, 96-108; Pallares, A., andFaus, V., Int'l. Endodon. J., 1995, 28, 266-269; Kouvas, V., Liolios,E., Vassiliadis, L., Parissis-Messismeris, S., and Boutsioukis, A.,Endod. Dent. Traumatol., 1998, 14, 191-195.

Additionally, the presence or absence of the smear layer is believed toplay an important role in the adhesive strength of a sealer to thedentinal walls. One study found a significant increase in adhesivestrength of AH26 sealer when the smear layer was removed. Gettleman, B.H., Messer, H. H., and ElDeeb, M. E., J. Endodon., 1991, 17, 15-20.These findings correlate with the results of another study demonstratingan increase in resistance to microleakage of AH26 when the smear layerwas removed. Economides, N., Liolios, E., Kolokuris, I., and Beltes, P.,J. Endodon., 1999, 25, 123-125.

Contrary to these findings, some studies have found that the presence orabsence of the smear layer has no significant effect on apical leakage.Evans, J. T. and Simon, J. H. S., J. Endodon., 1986, 12, 101-107.Kennedy, W. A., Walker, W. A., and Gough, R. W., J. Endodon., 1986, 12,21-27; Economides, N., Liolios, E., Kolokuris, I., and Beltes, P., J.Endodon., 1999, 25, 123-125; Timpawat, S., and Sripanaratanakul, S., J.Endodon., 1998, 24, 343-345.

It has been shown that removal of smear layer before sealing of the rootcanal system allows better adaptation between the obturation materialsand the root canal walls. Yamada, R. S., Armas, A., Goldman, M., andLin, P. S., J. Endodon., 1983, 9, 137-142; Czonstkowsky, M., Wilson, E.,and Holstein, F., Dental Clinics of N. Am., 1990, 34, 13-24. One studyexamined the adaptation of a mechanically softened gutta percha to thedentinal walls and reported that removal of the smear layer resulted inentry of gutta percha into the dentinal tubules. Pallares, A., and Faus,V., Int'l. Endodon. J., 1995, 28, 266-269. These authors reported nogutta percha penetration into the dentinal tubules in canals with intactsmear layer. Another study reported that when Thermafil, Ultrafill andcold lateral condensation techniques were used as obturation methods,all techniques showed significant resistance to microleakage with thesmear layer removed. Gencoglu, N., Samani, S., and Gunday, M., J.Endodon., 1993, 19, 558-562. Vertical condensation of gutta percha,Thermafil, and lateral compaction techniques with Ultrafill have alsobeen reported to reduce microleakage with the smear layer removed.Taylor, J. K., Jeansonne, B. G., and Lemon, R. R., J. Endodon., 1997,23, 508-512; Karagoz-Kucukay, I., and Bayirli, G., Int'l. Endod. J.,1994, 27, 87-93. In contrast to these findings, some studies havereported that removal of smear layer had no significant effect onmicroleakage of canals filled with laterally condensed gutta percha orThermafil and System B (warm vertical) obturation techniques. Saunders,W. P., and Saunders, E. M., J. Endodon., 1994, 20, 155-158; Kytridou,V., Gutmann, J. L., and Nunn, M. H., Int'l. Endodon. J., 1999, 32,464-474. Even if the smear layer can not be fully removed, one of skillin the art will recognize that it is desirable to remove as much of thesmear layer as possible, while sterilizing the portion that remains,prior to proceeding with filling, reconstruction, restoration, or finaltreatment.

The components of the smear layer are very small particles with a largesurface/mass ratio, which makes them very soluble in acids. Because ofthis characteristic, certain acids have been used in an attempt toremove the smear layer. Different formulations of ethylenediaminetetraacetic acid (EDTA) have been used to remove the smear layer fromthe surface of instrumented root canals, including REDTA (Roth EDTA).McComb, D., and Smith, D. C., J. Endodon., 1975, 1, 238-242. Someinvestigators, however, have questioned the effectiveness of REDTA byshowing that when used alone, REDTA removes the inorganic portion of thesmear layer but leaves an organic layer intact in the tubules. Goldman,M., Goldman, L. B., Cavaleri, R., Bogis, J., and Lin, P. S., J.Endodon., 1982, 8, 487-492. Sodium hypochlorite (NaOCl) has been shownto be very effective against this organic layer. When used alone, NaOClcan dissolve pulpal remnants, as well as predentin, but is ineffectivein removing the smear layer. The alternating use of EDTA and NaOCl,however, has been reported to be an effective method to remove the smearlayer. Goldman, M., Goldman, L. B., Cavaleri, R., Bogis, J., and Lin, P.S., J. Endodon., 1982, 8, 487-492; Yamada, R. S., Armas, A., Goldman,M., and Lin, P. S., J. Endodon., 1983, 9, 137-142; Baumgartner, J. C.,and Mader, C. L., J. Endodon., 1987, 13, 147-157. One study recommendsthe use of NaOCl during instrumentation, along with an EDTA rinsefollowed by a final flush with NaOCl. Baumgartner, J. C., and Mader, C.L., J. Endodon., 1987, 13, 147-157. Another study compared the abilityof various salts of EDTA to remove the smear layer and concluded thatall salts of EDTA were capable of removing the smear layer from thecoronal two thirds of root canals. In addition, the same study reportedthat tetrasodium salt, pH adjusted with HCl, is less expensive and justas effective as the more commonly used disodium EDTA. O'Connell, M. S.,Morgan, L. A., Beeler, W. J., and Baumgartner, J. C., J. Endodon., 2000,26, 739-743.

In 1993, a solution of EDTA and ethylenediamine was developed to work ina dual action. Aktener, B. O., and Bilkay, U., J. Endodon., 1993, 19,228-231. The goal was to see if a single irrigating solution can bedeveloped to remove the inorganic as well as the organic components ofthe smear. Many patent tubules were found, but more research was deemednecessary to determine the efficacy of this combination. Other studieshave added a quaternary ammonium bromide to EDTA to reduce its surfacetension. Goldberg, F., and Abramovich, A., J. Endodon., 1977, 3,101-105; Ciucchi, B., Khettabi, M., and Holz, J., Int'l. Endod. J.,1989, 22, 21-28. This addition increased the wetting effect on the canalwall and permitted deeper penetration of the solution intoirregularities. EDTAC, as it is named, was shown to be very effective insmear layer removal, reaching its peak effect at 15 minutes andincreasing the diameter of the opened dentinal tubules. Goldberg, F.,and Spielberg, C., Oral. Surg., 1982, 53, 74-77. Another study reportedeffective removal of the smear layer when using a solution of EDTA,carbamide peroxide, and propylene glycol. Tam, A., and Yu, D. C.,Compendium Cont. Ed. Dent., 2000, 21, 967-972. Recently, ethyleneglycol-bis (b-aminoethyl ether —NNNN-tetraacetic acid), EGTA, wasreported to be somewhat effective in removing the smear layer withoutinducing erosion commonly caused by EDTA. Calt, S., and Serper, A., J.Endodon., 2000, 26, 459-461.

The quantity of smear layer removed by an acid is directly related tothe concentration of the acid (pH) and the time of exposure. Morgan, L.A., and Baumgartner, J. C., Oral Surg. Oral Med. Oral Path., 1997, 84,74-78. Several studies used a 50% citric acid solution to treat canalwalls after instrumentation and found better penetration of rosin sealerinto the walls and improved adaptation of gutta percha when compared tountreated canals. Loel, D., J. A. D. A., 1975, 90, 148-151; Tidmarsh,B., J. Endodon., 1978, 4, 117-121; Baumgartner, J. C., Brown, C. M.,Mader, C. L., Peters, D. D., and Shulman, J. D., J. Endodon., 1984, 10,525-531. When citric acid was used as the sole agent for removal ofsmear layer, solutions at concentrations below 50% were ineffective.Yamada, R. S., Armas, A., Goldman, M., and Lin, P. S., J. Endodon.,1983, 9, 137-142; Takeda, F. H., Harashima, T., Kimura, Y., andMatsumoto, K., Int'l. Endodon. J., 1999, 32, 32-39. Lactic acid at 50%concentration is less effective than 50% citric acid for removal ofsmear layer. Wayman, B. E., Kopp, W. M., Pinero, G. J., and Lazzari, E.P., J. Endodon., 1979, 5, 258-265. This could possibly be attributed tothe viscosity of lactic acid. Additionally, alternating use of 10%citric acid and 2.5% NaOCl has also been reported to be a very effectivemethod for removing the smear layer. Wayman, B. E., Kopp, W. M., Pinero,G. J., and Lazzari, E. P., J. Endodon., 1979, 5, 258-265.

In 1989, one study reported that 25% tannic acid was effective inremoving the smear layer, but another study refuted these findings andexplained that tannic acid increased the cross-linking of exposedcollagen within the smear layer and within the matrix of the underlyingdentin, thus increasing organic cohesion to the tubules. Bitter, N. C.,Oral Surg. Oral Med. Oral Path., 1989, 67, 333-337; Sabbak, S. A., andHassanin, M. B., J. Prosthet. Dent., 1998, 79, 169-174.

Polyacrylic acid (Durelon liquid and Fuji II liquid) at 40% has beenreported to be very effective for removal of smear layer. Berry, B. A.,von der Lehr, W. N., and Herrin, B. K., J. A.D.A., 1987, 115, 65-67.Because of its potency, however, it is recommended that application ofPolyacrylic acid should not exceed 30 seconds.

Derivatives of oxine (8-hydroxy-quinoline) have been known to possessantiseptic qualities as early as 1895. Dequalinium compounds, whichbelong to this group, have been widely used in medicine againstinfections of bacteria, molds and fungi. Bis-dequalinium-acetate (BDA)has been shown to remove the smear layer throughout the canal, even inthe apical third. Kaufman, A. Y., Binderman, I., Tal, M., Gedalia, I.,and Peretz, G., Oral Surg., 1978, 46, 283-295; Kaufman, A. Y., OralSurg., 1981, 51, 434-441. BDA is well tolerated by the tissues withinthe periodontium and has a low surface tension that allows penetrationinto spaces that instruments cannot reach. BDA is also considered lesstoxic than NaOCl and can be used interoperatively as a root canaldressing. One study compared Salvizol (a commercial brand of 0.5% BDA)with 5.25% NaOCl and found both comparable in their ability to removeorganic debris, but only Salvizol was able to open dentinal tubules.Kaufman, A. Y., and Greenberg, I., Oral Surg., 1986, 62, 191-196.Another study reported Salvizol to be less effective at opening dentinaltubules compared to REDTA. Berg, M. S., Jacobsen, E. L., BeGole, E. A.,and Remeikis, N. A., J. Endodon., 1986, 12, 192-197.

The effects of the tetracycline family of antibiotics on removal ofsmear layer have also been studied to a degree. These materials havebeen used to demineralize dentin surfaces, uncover and widen theorifices of dentinal tubules and expose the dentin collagen matrix.These effects provide a matrix that stimulates fibroblast attachment andgrowth. Studies have shown that doxycyline HCl (100 mg/ml) is aneffective material to remove the smear layer from the surfaces ofinstrumented canals and those prepared for root-end filling materials.Barkhordar, R. A., Watanbe, L. G., Marshall, G. W., and Hussain, M. Z.,Oral Surg. Oral Med. Oral Path., 1997, 84, 420-423; Barkhordar, R. A.,and Russel, T., Cal. Dent. Assn. J., 1998, 26, 841-844; Haznedaroglu, F.and Ersev, H., J. Endodon., 2001, 27, 738-740. These studies speculatethat a reservoir of active antibacterial agent might be created sincedoxycycline readily attaches to dentin and can be readily releasedlater. Another study has reported increased demineralization effect whena 5% tetracycline/33% citric acid gel was used to treat teeth withmoderate periodontal disease. Jeong, S., Han, S., Lee, S., andMagnusson, I., J. Periodontol., 1994, 65, 840-847.

Apart from chemical solutions, mechanical methods, including ultrasonicinstrumentation, have been widely reported to be effective in removingthe smear layer from prepared tooth surfaces. Laser removal of the smearlayer has been shown to be successful as well for vaporizing tissues inthe main canal, removing the smear layer, and eliminating residualtissue in the apical portion of root canals. Since laser beams travel instraight lines, however, the use of lasers in curved canals is limited.

Smear layers are also formed when tooth material is removed preparatoryto restoration or other dental work, as it is for root canal situations.Moreover, in the restoration of bone, such as in orthopaedicrestorations, debris layers similar in many respects to endodontic smearlayers are also formed. It is now believed that their removal would behighly desirable as well.

Accordingly, it is believed to be highly desirable to remove the smearlayer from a prepared root canal space prior to filling the canal.However, removal of smear layer materials is very difficult toaccomplish. Moreover, there are no present methods likely to effectsubstantially complete removal of smear layers. Prior attempts have useda number of chemical species to remove the smear layer and sterilize theroot surface(s), but with indifferent results. The removal of smearlayer materials with a unitary solution to yield effective, convenient,and rapid smear layer removal is desired. All of this must beaccomplished without interfering with the essential purpose of rootcanal preparation or with the eventual restoration of the space. Removalof smear layers from tooth restoration sites, periodontal loci, andother prepared locations for dental and periodontic work is a furtherobject. Indeed, it is also believed to be desirable to remove smearlayers from orthopaedic and bone restoration sites within or without theoral cavity as well.

SUMMARY OF THE INVENTION

The present invention provides methods for removing smear layers fromand sterilizing endodontic excavations and other prepared tooth surfacesby irrigating with a mixture comprising disinfectant, detergent, andacid, especially organic acid. In a further aspect, the presentinvention relates to solutions for irrigating prepared tooth surfaces toremove smear layers as well as to restorations employing the method.Application to bone excavations is also contemplated.

It has now been discovered that a solution combining disinfectant,detergent, and acid is highly effective for removing the smear layer onprepared dental surfaces and dentinal tubules. Such solutions are usefulin a multitude of dental applications, including, but not limited to,root canal therapy; preparation of cavities; cosmetic and reconstructivedentistry such as caps, crowns, bridges, veneers, and the like; otherendodontic procedures; periodontic procedures; and bone preparation orrestoration. Such solutions are also useful in improving orthopaedicrestoration sites as well.

The term “smear layer” as used herein, is well known to persons skilledin the art of dentistry and refers to the complex accumulation oforganic and inorganic debris resulting from the mechanical preparationof a tooth surface. The smear layer comprises cutting debris, toothparticles, microorganisms, necrotic material, and other substancesresulting from preparation, and typically includes a superficial layeron the surface of a prepared tooth along with a layer or layers that arepacked into the adjacent dentinal tubules at varying depths up to about40 μm. In the context of orthopaedics, “smear layer” refers to similarlayers in prepared bone sites.

The term “disinfectant”, as used herein, refers collectively tocompositions that are able to suppress or eliminate bacterial or othermicroorganisms found in endodontic or periodontic sites. The term“disinfectant” includes antibiotics as that term is understood inpharmaceutical science.

The components of this invention comprise disinfectant, detergent, andacid. In a preferred embodiment, the disinfectant is an antibiotic. Itwill be apparent to one skilled in the art that the antibiotic should bestable in the acidic solutions of which it forms a part, should becompatible with the other components of the solution, and should retainits effectiveness for at least the time of preparation of the solutionand its application and residence time on or in the prepared tooth orbone surface. Examples of such antibiotics include, but are not limitedto, ansamycins, including rifamycins; cephalosporin; macrolides such asclarithromycin, josamycin, and oleandomycin; most polypeptides, such asbacitracin, capreomycin, enduracidin, enviomycin, gramicidin, mikamycin,ristocetin, thiostrepton, tyrocidine, viomycin, and virginiamycin; alltetracycline compounds, such as apicycline, chlortetracycline,clomocycline, demeclocycline, doxycycline, guamecycline, lymecycline,mecleocycline, methacycline, minocycline, oxytetracycline,penimepicycline, pipacycline, rolitetracycline, sancycline, mupirocin,and tetracycline-HCl; and tuberin. Most quinolones such asciprofloxacin, gatifloxacin, and moxifloxacin are not preferred, as theyare weak bases and have decreased effect in acidic solutions.Additionally, most B-lactam antibiotics, particularly penicillins, arealso not preferred, as they are generally unstable in acidic solutions.Exceptions, however, are amoxycillin, an acid-stable member of thepenicillin family, and similar compounds.

Tetracyclines are broad-spectrum antibiotics that are effective againsta wide range of microorganisms. They include tetracycline-HCl,minocycline, and doxycycline. Tetracyclines are bacteriostatic in natureand are generally more effective against gram-positive bacteria comparedto gram-negative bacteria. A reference to tetracycline shall be taken toinclude all members of the tetracycline family. A number of studies haveshown that tetracyclines significantly enhance healing after surgicalperiodontic therapy. Members of the family of tetracyclines arepreferred for use herein. Tetracyclines are preferred for a number ofreasons. One reason they are preferred is because they have many uniqueproperties along with their antimicrobial effect. For example,tetracycline-HCl has a low pH in concentrated solution and thus can actas a calcium chelator, and cause enamel and root surfacedemineralization. Tetracycline-HCl's surface demineralization of dentinis comparable to that seen using citric acid. In addition, it has beenshown that tetracycline-HCl is a sustentative medication and becomesabsorbed and released from tooth structures such as dentin and cementum.The use of tetracycline is also preferred because a very low portion ofthe general population exhibit allergies or other sensitivities totetracycline.

In another preferred embodiment, the disinfectant is an antimicrobialcompound. It will be apparent to one skilled in the art that theantimicrobial compound should be stable in the acidic solutions of whichit forms a part, should be compatible with the other components of thesolution, and should retain its effectiveness for at least the time ofpreparation of the solution and its application and residence time on orin the prepared tooth or bone surface. Examples of such antimicrobialcompounds include, but are not limited to, chlorhexidine compounds.Chlorhexidine gluconate is preferred. One example of a suitablechlorhexidine gluconate solution is a commercially available 0.12%solution known as “peridex”. Additionally, the use of chlorhexidinegluconate has been found to be especially desirable in patients whoexhibit sensitivity or allergy to tetracycline compounds.

It will also be recognized by one skilled in the art that the detergentused should be stable in acidic solution with an antibiotic compound.Additionally preferable is a detergent that reduces surface tension ofthe solution, thus providing an increased wetting effect and permittingenhanced penetration of the irrigation solution into dentinal tubulesand irregular spaces that are otherwise difficult to reach. Furthermore,the detergent should be one suitable for use in situ in dentalapplications without deleterious effect to the human or animal subject.

In a preferred embodiment, the detergent is a non-ionic surfactant orsimilar compound, preferably one commonly used in the food and drugindustry or approved for use by the Food and Drug Administration.Examples of such compounds include, but are not limited to, mono- anddi-glycerides; sucrose esters; sorbitan esters (also known as SPANs),particularly sorbitan monostearate; sorbitols; polysorbates(polyoxyethylene sorbitan esters, also known in industry as TWEENs),particularly polysorbate 20, polysorbate 60, polysorbate 65, andpolysorbate 80; stearoly lactylates; lecithin and derivatives;polyglycol fatty acid esters; p-Cymene; quaternary ammonium compounds;sodium alkyl sulfonates; triethanolamine; and alkyl polysaccharides.

In another preferred embodiment, the detergent used is selected from thegroup of sorbitan esters or polysorbates. One exemplary member of thepreferred class is polysorbate 80 (polyoxyethylene sorbitan monooleate).

It will also be apparent to one of skill in the art of dentistry thatthe acid used should be suitable for dental application. Thus, the acidshould be nontoxic in the applicable concentration and amount used inthe irrigation process and should also be compatible with the detergentand disinfectant selected as the other components of the solution.Preferred acids must also be capable of dissolving the organic andinorganic components of the smear layer within the chosen exposure time,but without inducing unwanted erosion of the tooth and surroundingsurfaces.

In another preferred embodiment, the acid is an organic acid, preferablyhaving pKa values between 1.5 and 5. Further preferred are carboxylicacids or other acids with a polar nature and pKa values between 2 and 5.In a further preferred mode of the present invention, an acid with a pKavalue between about 2.75 and 3.75 is used. One exemplary member of thepreferred class is citric acid. Citric acid is particularly suitablewhen tetracycline is chosen as the disinfectant, because citric aciddoes not diminish or otherwise alter the antibacterial effect oftetracycline.

It will be apparent to one skilled in the art, however, that strongeracids may also be preferred for use in the present invention providedthat the time of application of the solution is shortened accordingly.As such, stronger acids including, but not limited to, chloracetic,maleic, saccharic, tartaric, and polyacrylic may be used, having pKavalues ranging from about 0.5 to about 3.0. Mixtures may also be used.In some embodiments inorganic acid, specifically phosphoric acid mayfind utility so long as the essential properties of the solution aremaintained.

The disinfectants are present in the solutions of the present inventionin weight percentages of from about 1 to about 5 percent of the solutionand preferably in amounts of from about 2 to about 4 weight percent,with amounts of about 3 percent being even more preferred, especiallywhen the disinfectant is a tetracycline.

The detergent is preferably present in the solutions of the invention inweight percentages of from about 0.1 to about 1.5 percent of thesolution, with amounts of from about 0.25 to about 1.0 percent beingmore preferred. Amounts by weight of about 0.5 percent are generallymost preferred depending upon the detergent, especially when thedetergent is a polysorbate.

The acids of the invention are present in the solutions in amounts offrom about 0.5 to about 10 percent by weight of the solution, preferablyfrom about 3 to about 6 percent. More preferred are solutions havingweight percentages of acid, especially organic acid, of from about 4 toabout 5 percent.

In general, the solutions of the invention are aqueous and watercomprises the bulk of the balance of the composition. Solutions of theinvention may also include other compounds, however, so long as they donot interfere with the essential functions of the principal components,do not cause them to degrade and do not interfere with the convenienceand utility thereof. Such additional additives may include colorants,flavorants, stabilizers, and other materials conventionally added todental or orthopaedic solutions. One particularly useful adjuvant may bechelating agents capable of rendering chelatable materials, especiallymetals, soluble. Indeed, use of a polyfunctional acid may achieve thisgoal. It will be recognized by one of skill in the art that regardlessof the components or additives in the solution, the resulting solutionshould be sterile so that the objectives of the invention are achieved.In all cases, such materials are present in effective amounts toaccomplish their objectives.

In a preferred embodiment of the current invention, the solutioncomprises an aqueous solution of 3% doxycycline, 0.5% polysorbate 80,and 4.25% citric acid by weight. While these components have previouslybeen used separately and in high concentrations in efforts to remove thesmear layer, the three components as described above have not beencombined as in the present invention. Additionally, studies performed inconjunction with the present invention using a solution of 3%doxycycline, 0.5% polysorbate 80, and 4.25% citric acid have shown lowlevels of cytotoxicity and no mutagenicity when compared to all purposebleach, which had previously been used to disinfect tooth preparations.

Additional studies performed with a solution of doxycycline, polysorbate80, and citric acid have shown further superior properties. For example,the solution exhibits lower toxicity than eugenol, 3% H₂O₂, Ca(OH)₂paste, peridex, and EDTA, and exhibits greater antimicrobial propertiesthan 5.25% NaOCl, even when applied to infected root canals for briefperiods of time, such as 2 and 5 minute intervals of use. It will beapparent to those of skill in the art that a solution such as the onedescribed herein (i.e., one that has improved biocompatibility and iseffective in a shorter period of time than previously known treatments)is highly desirable, particularly in the health care fields. Studieshave also shown that, unlike other previously known dental irrigants,the solution of the present invention maintains the desiredantimicrobial effect at dilutions up to 1:200. In contrast, thebactericidal properties of NaOCl cease to exist at dilutions of about1:32, and EDTA is ineffective in anything other than an undiluted state.In a further study undertaken to evaluate the antimicrobial propertiesof the presently described solution, human root canals were infectedwith whole saliva and then treated with either 5.25% NaOCl or a solutionof doxycycline, polysorbate 80, and citric acid. While the NaOCleffectively disinfected only 37 out of 60 teeth, the solution of thepresent invention disinfected 59 out of 60 teeth. The importance anddesirability of such improved antimicrobial properties can be easilyappreciated, particularly by those of skill in the art.

The present invention is directed to methods for sterilizing andremoving the smear layer on a prepared tooth or canal surface comprisingirrigating the surface with a solution comprising disinfectant,detergent, and acid. In preferred modes of the invention, thedisinfectant is an antibiotic that is sufficiently stable in an acidicenvironment. It is further preferred that the antibiotic be atetracycline compound. In a further preferred embodiment, thetetracycline compound is doxycycline. In other preferred modes of theinvention, the detergent is an FDA-approved additive, preferably apolysorbate or sorbitan ester compound. In another preferred mode of theinvention, the detergent is polyoxyethylene sorbitan monooleate(polysorbate 80).

In another preferred aspect of the present invention, the acid is anorganic acid, preferably having a pKa between 1.5 and 5. In a furtherpreferred embodiment, the organic acid has a pKa between 2 and 4;preferably between 2.75 and 3.75, such as that of citric acid. In afurther embodiment, the acid is phosphoric acid.

The methods of the present invention can be used on surfaces ofinstrumented root canals, sites prepared for periodontic procedures,sites prepared for tooth restoration or reconstruction, and sitesprepared for bone restoration or reconstruction. In a preferred mode ofthe present invention, the prepared tooth surface is irrigated forbetween 1 minute and 1 hour, preferably between 1 and 30 minutes andmore preferably from about 1 to about 10 minutes.

Although the uses described above are exemplary for the presentinvention, there are other embodiments that may be foreseen by thoseskilled in the art. The solution of the present invention can also haveuse in preparation for implants in the animal body. Such foreseeablepreparations include use with cochlear, cranial, sternum, other customimplants or functional shapes made for the body. Other embodiments canbe used for preparation for insertion of universal plates for orthopedicuse, bone screws, rods & pins for orthopedic use (IM nails, femoral rodsor plugs, long bone fractures, etc.), tendon anchors, suture anchors andtacks, graft retainers and marrow sampling ports.

For use in connection with removal of smear layer from bonypreparations, either in the mouth or upon skeletal bone, a prepared siteis irrigated for from 1 minute to one hour, preferably from 1 minute toabout 30 minutes, with from about 1 to 10 minutes being preferred. By“irrigation” is meant contacting the site with the solution. It ispreferred to provide a flow of such solution over the surfaces of thesite, however, this need not be performed continuously. Flow of solutionmay be accompanied by air entrainment to assist in smear layer removalthrough action of the ensuing bubbles. Other physical means of assistingwith smear layer removal may accompany irrigation and all such areencompassed hereby.

Following irrigation, the site is dried and used for the intendedrestoration.

It has further been discovered that the solutions of the presentinvention can be particularly effective when used following an initialrinse comprising NaOCl. In such applications, NaOCl may be used as anirrigant in conjunction with instrumentation of a surface. Followinginstrumentation and use of the NaOCl rinse, a final rinse ofdisinfectant/detergent/acid solution may be applied to the surface. Thismethod is believed to have an improved effect on smear layer removalbecause the initial NaOCl rinse removes some organic materials from thesmear layer, while the disinfectant/detergent/acid solution removesinorganic and residual organic materials. When NaOCl is used inconjunction with the solution of the present invention, it is preferredthat the concentration of NaOCl be between about 1% and about 6% byweight, and the concentration is most preferably between about 1.3% andabout 5.25% by weight. Because testing indicates that there is nosignificant difference in performance within this concentration range,and high concentrations of NaOCl are known to be more toxic than lowerconcentrations, it is recommended that NaOCl at the lower end of thegiven concentration range be used. A method comprising the use of aninitial rinse of a solution of 1.3% NaOCl by weight followed by a finalrinse of a solution comprising doxycycline, polysorbate 80, and citricacid is particularly desirable.

In a further embodiment, it may be beneficial to take steps to increasethe shelf life of the prepared solution. For example, in a preferredembodiment, the prepared solution is dehydrated and then the resultingpowder is rehydrated with an appropriate amount of distilled water priorto use. Persons of skill in the art will recognize that there are manyways to effectively dehydrate the solution, for example by dessicating,lyophilizing, spray drying, or by any other method that renders thesolution in a shelf-stable powdered form. By this method, the shelf lifeof the solution is increased significantly. Furthermore, the powderedsolution may simply be rehydrated with an appropriate amount ofdistilled water and then used to irrigate prepared surfaces inaccordance with the procedures described above.

EXAMPLES

The invention is illustrated by the following examples, which are notintended to be limiting.

Example 1 Removal of Smear Layer from Root Canal Walls

Extracted maxillary and mandibular human teeth were used for this study.The working lengths of these teeth were between 21-25 millimeters. Theteeth were scaled of any calculus and other surface debris (soft tissueand/or alveolar bone) using hand scalers. After preparing conventionalaccess preparations through the incisal or occlusal surfaces of the testteeth, a K-type file (size 10) was used to determine the working lengthof each tooth by penetrating the apical foramen and pulling back to theclinical apical foramen. A combination of passive step back techniqueand rotary files (Rivera and Walton 2002) were used to clean and shapethe root canals. Each canal was cleaned and shaped using a combinationof passive step back and Rotary 0.04 Taper NITI files (Rivera andWalton, 2002). The apex of each tooth was enlarged to size 30 files.

The teeth were randomly assigned to one of the two groups: Group 1:Non-surgical endodontic therapy (NSET) was performed using 5.25% sodiumhypochlorite as an irrigant. After complete cleaning and shaping, thecanals were irrigated with 1 ml of NaOCl and a barbed broach wrappedwith cotton was taken to the canal terminus and left for five minutes toensure uniform direct contact of the irrigant with the entire canal.Upon removal of the barbed broach, the canal was irrigated with 4 ml ofNaOCl and rinsed with 10 ml of distilled water. Group 2: NSET wasperformed using 1% sodium hypochlorite as root canal irrigant. Aftercomplete cleaning and shaping of teeth in this group, the canals wereirrigated with 1 ml of a mixture of 3% doxycycline, 0.5% polysorbate 80,and 4.25% citric acid, hereinafter referred to as “ADD” (acid,disinfectant, and detergent) solution, and a barbed broach wrapped withcotton was taken to the canal terminus and left for five minutes toensure uniform direct contact of the irrigant with the entire canal.Upon removal of the barbed broach, the canal was irrigated with 4 ml ofADD and rinsed with 10 ml of distilled water. After irrigation, theteeth were split in half using a diamond saw and constant water spray.Half of each tooth was placed into a gluteraldehyde solution for 24 h.The fixed specimens were then rinsed twice by a sodium buffered solution(pH 7.2), treated with osmium tetraoxide for one hour, rinsed withascending concentrations of ethyl alcohol 30%-100%, and then placed in adessicator for 24 h. Finally, each specimen was mounted on a specialbutton, and coated with 25 μm of gold-palladium (Au—Pd) (Wakabayashi etal., 1995). The presence or absence of smear layer at the coronal,middle, and apical portion of each canal was evaluated. The erosion ofdentinal tubules was also assessed at different levels of each canal.

Examinations of the specimens showed presence of smear layer on theentire root canal walls of all teeth prepared in Group 1. In contrast,the walls of canals in Group 2 had no detectable smear layer in anysample. There was a significant difference between the two groups. Noerosion was noted in the dentinal tubules at various levels of eachcanal treated with ADD.

Example 2 Removal of Smear Layer from Coronal Cavity Preparations

Sound third molars were collected and stored in deionized water. Class Ipreparations were made following conventionally accepted procedures inthe occlusal surfaces of the teeth. The teeth were randomly divided intotwo groups.

Group 1: A solution of 5.25% of NaOCl was left in the cavity for 5minutes. After this treatment each preparation was rinsed with copiousamounts of water to eliminate the residual effect of sodiumhypochlorite.

Group 2: ADD solution was left in the cavity for 5 minutes. After thistreatment each preparation was rinsed with copious amounts of distilledwater to eliminate the residual effect of ADD.

After irrigation, the crowns were split in half using a diamond saw andconstant water spray. Half of each sample was placed into agluteraldehyde solution for 24 h. The fixed specimens were then rinsedtwice by a sodium buffered solution (pH 7.2), treated with osmiumtetraoxide for one hour, rinsed with ascending concentrations of ethylalcohol 30%-100%, and then placed in a dessicator for 24 h. Finally,each specimen was mounted on a special button, and coated with 25 μm ofgold-palladium (Au—Pd) (Wakabayashi et al., 1995). The presence orabsence of smear layer on the walls of each cavity was evaluated. Theerosion of dentinal tubules was also assessed.

Examinations of the specimens showed presence of smear layer on thewalls of all teeth prepared in Group 1. In contrast, the walls ofcavities in all samples in Group 2 had no smear layer and had patentdentinal tubules. There was a significant difference between the twogroups.

No erosion was noted in the dentinal tubules of cavities in Group 2treated with ADD.

Example 3 Removal of Smear Layer from Prepared Crown Preparations

Sound third molars were collected and stored in deionized water. Crownpreparations were made following conventionally accepted procedures. Thesamples were randomly divided into two groups.

Group 1: A cotton pellet saturated with a 5.25% NaOCl solution was lefton the crown preparations for 5 minutes. After this treatment eachpreparation was rinsed with copious amounts of distilled water toeliminate the residual effect of sodium hypochlorite.

Group 2: A cotton pellet saturated with ADD solution was left on thecrown preparations for 5 minutes. After this treatment each preparationwas rinsed with copious amounts of distilled water to eliminate residualADD.

After irrigation, the crown preparations were split in half using adiamond saw and constant water spray. Half of each sample was placedinto a gluteraldehyde solution for 24 h. The fixed specimens were thenrinsed twice by a sodium buffered solution (pH 7.2), treated with osmiumtetraoxide for one hour, rinsed with ascending concentrations of ethylalcohol 30%-100%, and then placed in a dessicator for 24 h. Finally,each specimen was mounted on a special button, and coated with 25 μm ofgold-palladium (Au—Pd) (Wakabayashi et al., 1995). The presence orabsence of smear layer on the walls of each cavity was evaluated. Theerosion of dentinal tubules was also assessed.

Examinations of the specimens showed presence of smear layer on thesurface of all crowns prepared in Group 1. In contrast, the surfaces ofcrowns prepared in Group 2 had no smear layer and had patent dentinaltubules. There was a significant difference between the two groups. Noerosion was noted in the dentinal tubules of crown preparations in Group2.

Example 4 Removal of Smear Layer from Root Surfaces of Teeth

Sound extracted teeth were used in this experiment. The coronalone-third root surfaces of these teeth were curetted followingconventionally accepted procedures. The samples were randomly dividedinto two groups.

Group 1: A cotton pellet saturated with a 5.25% NaOCl solution was lefton the surfaces of root preparations for 5 minutes. After this treatmenteach root surface was rinsed with copious amounts of distilled water toeliminate the residual effect of sodium hypochlorite.

Group 2: A cotton pellet saturated with ADD solution was left on theroot surface preparations for 5 minutes. After this treatment eachpreparation was rinsed with copious amounts of distilled water toeliminate the residual effect of ADD.

After irrigation, the entire tooth was split in half using a diamond sawand constant water spray. Half of each sample was placed into agluteraldehyde solution for 24 h. The fixed specimens were then rinsedtwice by a sodium buffered solution (pH 7.2), treated with osmiumtetraoxide for one hour, rinsed with ascending concentrations of ethylalcohol 30%-100%, and then placed in a dessicator for 24 h. Finally,each specimen was mounted on a special button, and coated with 25 μm ofgold-palladium (Au—Pd) (Wakabayashi et al., 1995). The presence orabsence of smear layer on the root surfaces was evaluated. The erosionof dentinal tubules was also assessed.

Examinations of the specimens showed presence of smear layer on thesurface of all roots prepared in Group 1. In contrast, the surfaces ofroots prepared in Group 2 had no smear layer and had patent dentinaltubules. There was a significant difference between the two groups. Noerosion was noted in the dentinal tubules of root preparations in Group2.

Example 5 Removal of Smear Layer from Root End Cavity Preparations

Extracted, human single-rooted teeth were used in this part of theexperiment. The clinical crown of each tooth was removed at thecemento-enamel junction using a #701 fissure bur in a high-speedhandpiece and water spray.

The working length of each canal was determined by placing and moving a#15 K file apically in the canal until it exited from the apicalforamen. After enlarging the apical foramen to a #40 K file, the rest ofthe canal was cleaned and shaped using a combination of passivestep-back technique and rotary instruments (Rivera and Walton, 2002),while a 5.25% NaOCl solution was used as the intracanal irrigant.

The instrumented canals were dried with paper points and obturated withlaterally condensed gutta-percha and Roth 811 sealer. The accesscavities were closed with Cavit. The roots were then wrapped in moistgauze and stored in a closed glass bottle at room temperature and 100%humidity for one week.

Two coats of nail polish were applied to the external surface of eachroot. Apical root resections were then performed by removing 3-4 mm ofthe apex, at a 90° angle to the long axis of the root, with a #701fissure bur in a high-speed hand piece with water coolant.

Apical cavity preparations were made in each of the roots. A #1 roundbur in a high-speed handpiece with water coolant was used to create asmall opening into the gutta-percha filling material. The cavities wereenlarged and deepened to approximately 3 mm using a #701 fissure bur inhigh-speed handpiece with water spray. A # 541 med 108/010: HIDI diamondbur in a high speed handpiece with water spray was then used tostandardize the preparation to a diameter of 1.5 mm and a depth of 3 mm.

The roots were then randomly divided into two groups. In Group 1, theapical preparations were rinsed 5.25% NaOCl. The solution was left inthe cavity for five minutes and then rinsed with 10 ml of distilledwater. In Group 2, the preparations were rinsed with 5 ml of ADD. Thissolution was also left in the root end cavities for five minutes andthen rinsed with 5 ml of distilled water. After drying with paperpoints, the roots were split in halves using a slow speed diamond saw(Labut Agar Scientific, Cambridge, England).

Half of each sample was placed into a gluteraldehyde solution for 24 h.The fixed specimens were then rinsed twice by a sodium buffered solution(pH 7.2), treated with osmium tetraoxide for one hour, rinsed withascending concentrations of ethyl alcohol 30%-100%, and then placed in adessicator for 24 h. Finally, each specimen was mounted on a specialbutton, and coated with 25 μm of gold-palladium (Au—Pd) (Wakabayashi etal., 1995). The presence or absence of smear layer on the root endcavity preparations was evaluated. The erosion of dentinal tubules wasalso assessed.

Examinations of the specimens showed presence of smear layer on thesurface of all cavity preparations prepared in Group 1. In contrast, thesurfaces of cavities prepared in Group 2 had no smear layer and hadpatent dentinal tubules. There was a significant difference between thetwo groups. No erosion was noted in the dentinal tubules of root endcavity preparations in Group 2.

Example 6 Removal of Smear Layer from Prepared Bone Sites

A 0.5 mm dental drill was used to drill cavities in extracted tibiasegments of mature male rats. 5 ml of distilled water was used as anirrigant during drilling. The resulting cavities were 0.6 to 1.0 mm indiameter.

Following drilling, each prepared cavity was irrigated with 5 ml of asolution of 3% tetracycline, 0.5% polysorbate 80, and 4.25% citric acid.After irrigation, the solution was left in the bone cavity for 5minutes. The bones were then split in half using a diamond saw andconstant water spray. Half of each bone fragment was then placed in 4%formaldehyde solution for 24 hours. The segments were then dehydrated inserial ethanol concentrations and embedded in polymethylmethacrylate(PMMA). The specimens were then coated with a gold palladium film forSEM examination.

Visual SEM inspection showed substantially complete removal of the smearlayer on all samples.

Example 7 Removal of Smear Layer in Conjunction with NaOCl Rinse

Eighty extracted maxillary and mandibular single and multi-rooted humanteeth were used for this study. In multi-rooted teeth, the root with thelargest canal was included in the study. Teeth with previous root canaltreatment were excluded. The teeth were randomly divided into 7experimental groups of 10 teeth each and two control groups of 5 teetheach. The teeth were grouped according to the type of irrigants andfinal rinses used during and after instrumentation.

After preparing a conventional access preparation for each tooth, aK-type file (size 10 or 15) was used to determine the working length bypenetrating the apical foramen and pulling back to into the clinicalapical foramen. The working length of each tooth was between 21 and 25millimeters. Each canal was instrumented using a combination of passivestep back and Rotary 0.04 Taper NITI files (Millefer ProFile,Switzerland). The apical foramen of each tooth was enlarged to a size 30file. Distilled water, various concentrations of NaOCl (0.65%, 1.3%,2.6%, and 5.25%), and ADD solution were used as intracanal irrigants.One milliliter of irrigation solution was used to irrigate the rootcanal between each hand and rotary instrument. A total of 10 mL ofirrigant was used in each canal. The irrigants were delivered with a27-gauge plastic needle (Ultra dent Products South Jordan, Utah, USA)that penetrated to within 1-2 mm from the working length in each canal.Each canal was filled with an irrigant during instrumentation. Theinstrumentation time for each root canal was approximately 18-20minutes. To determine the effect of control and experimental solutionsas a final rinse on the surface of instrumented root canals, the canalswere then treated with 5 mL of one of the following solutions for 2minutes: sterile distilled water (positive control), 17% EDTA (negativecontrol), 5.25% NaOCl, and ADD solution.

After instrumentation, each canal was initially irrigated with 1 mL ofone of the above solutions. After 2 minutes, each canal was irrigatedwith 4 mL of one of the control or experimental solutions as a finalrinse. These irrigants were also delivered with a 27-gauge plasticneedle (Ultra dent Products South Jordan, Utah, USA) that penetrated towithin 1-2 mm from the working length in each canal. The total exposuretime for the final rinse was approximately 2 minutes. The canals werethen irrigated with 10 mL of sterile distilled water and dried withpaper points. The teeth were then split longitudinally and half of eachtooth was placed in a 2% glutaraldehyde solution for 24 hours. The fixedspecimens were rinsed 3 times with a sodium cacodylate buffered solution(0.1 M, pH 7.2), incubated in osmium tetraoxide for 1 hour, dehydratedwith ascending concentrations of ethyl alcohol (30%-100%), and thenplaced in a dessicator for at least 24 hours. Each specimen was mountedon an aluminum stub, and coated with 25 μm of gold-palladium andexamined under a scanning electron microscope.

The specimens were examined to determine the extent of both removal ofsmear layer and erosion of the dentinal tubules. The most effectivecombination at removing smear layer while minimizing erosion was 5.25%NaOCl-ADD (expressed as “initial rinse solution-final rinse solution”),followed by 2.6% NaOCl-ADD, 1.3% NaOCl-ADD, ADD-ADD, distilledwater-ADD, 5.25% NaOCl-5.25% NaOCl, and distilled water-distilled water(the least effective combination). A combination of 5.25% NaOCl-17% EDTAwas effective for removing smear layer, but resulted in moderate tosevere erosion of the dentinal tubules. Statistically, there was nosignificant difference between the performance of initial rinses of1.3%, 2.6% and 5.25% NaOCl in combination with a final rinse of ADD.

Example 8 Antibacterial Effect of Varying Dilutions of ADD Solution

Two methods were used to determine the extent of antimicrobial activityof ADD in comparison with NaOCl and EDTA at varying dilutions. In thefirst test, the zone of inhibition was measured on plates inoculatedwith Enterococcus faecalis. An overnight culture of E. faecalis (ATCC4082) was standardized to 0.11 optical density (O.D.) measured at 570nm. One hundred microliters of the microorganism was spread onto atrypticase soy agar (TSA) plate with the use of a sterile L-shaped glassrod. One-quarter inch sterile S & S filter paper (Schleicher & Schuell)was placed in each of the four quadrants of the TSA plate. Twentymicroliters of sterile saline, 5.25% NaOCl, ADD, or 17% EDTA was addedonto the filter papers. Eight replicates were prepared for each of thesample solutions. The plates were incubated overnight at 37° C. for 24hours and the zones of inhibition were measured in millimeters.

In the second test, the minimum inhibitory concentration (MIC) wasmeasured. E. faecalis was cultured overnight and the concentration wasadjusted to 0.11 O.D. at 570 nm. The test solutions were seriallydiluted from 1:2 up to 1:2048 dilutions. One ml of 2× trypticase soybroth (TSB) and the same amount of a test solution were mixed in varioustubes. One hundred microliters of the standardized E. faecalis was addedinto each of the test tubes and were incubated overnight at 37° C. Thepresence or absence of turbidity was determined the following day.

In order to determine whether the test solutions had inhibitory orbactericidal effect, a high concentration of bacteria (1×10⁸ c.f.u. ofE. faecalis) was exposed to 2 ml of undiluted and 1:2 dilution ADD orNaOCl for 2 or 5 minutes. Samples were placed on TSA media to determineviability of any remaining bacteria.

Dilution of the solutions reduced the zones of inhibition for allsolutions, albeit at different rates. NaOCl and ADD were both moreeffective when undiluted than at 1:5 or 1:10 dilution, but both stillhad some bactericidal effect upon dilution. EDTA, however, demonstratedno bactericidal properties when diluted five- or ten-fold. With respectto the minimum inhibitory concentration tests, EDTA exhibited noantibacterial effects at any dilution level, while NaOCl was effectiveat dilutions up to 1:32 and ADD was effective at dilutions up to 1:200.Additionally, after both 2 and 5 minutes of exposure at both undilutedand 1:2 dilution levels, ADD resulted in completely negative cultures,while NaOCl did not.

Each of the patents, publications, and other documents mentioned orreferred to in this specification is herein incorporated by reference inits entirety. Those skilled in the art will appreciate that numerouschanges and modifications may be made to the preferred embodiments ofthe invention and that such changes and modifications may be madewithout departing from the spirit of the invention. It is thereforeintended that the appended claims cover all such equivalent variationsas fall within the true spirit and scope of the invention.

1. A method for removing smear layer from a prepared tooth surface comprising irrigating the surface with a sterile solution comprising about 3% doxycycline by weight of the solution, about 0.5% polysorbate 80 by weight of the solution, and about 4.25% citric acid by weight of the solution.
 2. A method for removing smear layer from a prepared tooth comprising the steps of: (a) irrigating the surface with a solution of 1.3% NaOCl by weight of the solution; and (b) further irrigating the surface for from about 2 to about 5 minutes with a sterile solution comprising about 3% doxycycline by weight of the solution, about 0.5% polysorbate 80 by weight of the solution, and about 4.25% citric acid by weight of the solution.
 3. The method of claim 2 wherein the solution of step (b) is diluted up to about 1:200.
 4. The method of claim 2 wherein the components of the solution of step (b) are provided in powdered form, further comprising the step of rehydrating the powder with distilled water before irrigation. 